The high speed supernatant of Novikoff hepatoma cells contains factors which are capable of increasing the rate of DNA synthesis by the homogeneous beta-polymerase. These factors have been purified to varying degrees: Factor I is about 20% homogeneous, Factor IId is 90% homogeneous, and Factors III and IV are pure. The proposed research is aimed at 1) identifying any and all factors participating in or regulating DNA synthesis; 2) purifying each factor to homogeneity; 3) characterizing each factor with regard to enzymatic activity, molecular weight and subunit structure, stoichiometry of its action on in vitro DNA synthesis, binding to DNA or DNA polymerase, interaction with other factors, subcellular localization and amounts present under different physiological conditions; 4) clearly establishing a role for each in DNA replication and/or repair in vivo. The beta-polymerase as extracted from Novikoff hepatoma cells exists as an aggregate of proteins sedimenting at 7.3S. The homogeneous enzyme sediments at 3.3S, and intermediates of 4.1S and 5.8S have been identified. Factor IV has been shown to bind to the 3.3S enzyme in a 1:1 ratio to form the 4.1S intermediate. The other proteins needed to form the 5.8S and 7.3S forms are being sought by the use of bifunctional cross-linking agents and attempting to isolate the complex as a unit. Preparation of antibody to the polymerase and factor III is in progress and will be used to determine the time of synthesis in the cell cycle, amount present in normal vs. tumor cells, and subcellular localization using radiolabelled and fluorescent antibody techniques. BIBLIOGRAPHIC REFERENCES: D.W. Mosbaugh, D.M. Stalker, G.S. Probst and R.R. Meyer (1977): Novikoff hepatoma deoxyribonucleic acid polymerase. Identification of a stimulatory protein bound to the beta-polymerase. Biochemistry, 16, (in press). D.M. Stalker and R.R. Meyer (1977): Novikoff hepatoma deoxyribonucleic acid polymerase. Superhelically coiled, closed circular DNA as template for the beta-polymerase. (Manuscript in preparation, to be submitted to Proc. Nat. Acad. Sci.).